Production and differentiation of a vascular graft grown in the host’s peritoneal cavity: devices and bioreactors

Peter Stickler (2009). Production and differentiation of a vascular graft grown in the host’s peritoneal cavity: devices and bioreactors MPhil Thesis, AIBN Australian Institute for Bioengineering and Nanotechnology, The University of Queensland.

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Author Peter Stickler
Thesis Title Production and differentiation of a vascular graft grown in the host’s peritoneal cavity: devices and bioreactors
School, Centre or Institute AIBN Australian Institute for Bioengineering and Nanotechnology
Institution The University of Queensland
Publication date 2009-01
Thesis type MPhil Thesis
Supervisor Prof. Julie Campbel
Prof. Darren Martin
Total pages 249
Total colour pages 63
Total black and white pages 186
Subjects 06 Biological Sciences
Abstract/Summary The main question that this thesis addresses is what is the optimal way of producing tissue grown in the peritoneal cavity around a foreign body for its use as a vascular graft? It is known that a foreign body implanted into the peritoneal cavity induces an inflammatory response with cells recruited from within the peritoneal cavity to encapsulate the foreign body. Over the course of two to three weeks these cells produce an organised matrix and differentiate to become myofibroblasts. Tubes of these ‗tissue capsules‘ have been transplanted into the arterial vasculature in several animal models where the tissue capsule differentiates into an arterial structure. This structure consists of a layer of smooth muscle-like cells, adventitia of dense connective tissue including vasa-vasorum and an endothelial layer of flattened mesothelial cells. In order to determine whether the tissue would further differentiate ex vivo in response to mechanical stimulus an in-vitro bioreactor system was built to house tissue capsules produced in a variety of animal models. This bioreactor system could house 4 tissue capsules under physiological conditions including standard pulse rates, pressures and temperatures experienced by an artery. Boiled blood clot (BBC) scaffolds were implanted into the peritoneal cavity of rats to produce tissue capsules. After two weeks of development in the peritoneal cavity, tissue capsules were harvested and implanted into the bioreactor. Tissue capsules grafted into the bioreactor were subjected to mechanical force for a range of time-points, pressure, pulse and flow rates. When analysing tissue immunohistochemically, elastin, myosin, αSMA and desmin were detected. This staining was not consistent across all samples and only present in small parts of some tissue tested. Western analysis did not show any expression of αSMA or myosin. Finally the morphology of the tissue also resembled that of tissue previously implanted into the arterial circulation, but development of mechanical properties were not to the extent that would make the tissue useful as a vascular graft. The bioreactor system was thus modified to be able to house tissue for a period of 3 weeks. This system successfully housed tissue capsules under mechanical force in physiological ranges. Next, a range of materials were tested for their ability to be included into the peritoneal implant device used for the large animal model. Elasteon 80A did not produce any cellular growth or peritoneal pathology in all implanted samples (n = 4). Cloisite, a pro-inflammatory material produced large tissue capsule development over a 2 week implant period in 25% of samples however this tissue was heavily adhered to the greater omentum and dependent on its vascular supply. This data suggested that Elasteon could be used to coat the outer surface of a peritoneal implant device to decrease the rate of peritoneal adhesions. Three devices were designed and fabricated for their use in generating tissue for the modified Mitrofanoff procedure which requires a length of tissue to be implanted between the umbilicus and the bladder as a fistula. In all three cases no implantable material was produced that could be used for this procedure. To modify the device that could be used to produce tissue for any surgical application, a range of devices was produced and the animal model was changed to pigs. Materials incorporated into these devices include Dexon mesh and polyethylene. These devices also did not produce any tissue that could possibly be used as a vascular graft. A novel material, polymer BD347 was then produced for use in developing tissue within the interior of the device to provide greater growth and mechanical properties for developing a vascular graft. In toxicological studies, the replacement rate of cells was unaffected after seven days of incubation of fibroblasts at confluence with the polymer. A range of mechanical properties from pig vasculature was gained so that a sheet of polymer with similar properties to that of a vascular graft could be made. This polymer was fabricated as a tube and implanted into the peritoneal cavity of rats. The implanted polymer remained free-floating with a capsule of tissue in 78% of cases. A device was designed that has the ability to impart a physiological pulsation force on the developing tissue capsule in the peritoneal cavity using a sheep model. When two devices were implanted for a period of 10 days in each animal these devices produced no complications for the animal. Upon harvest all devices were free of adhesion and did not cause any peritoneal or dermal infection. In 100% of cases this device produced tissue that was thick and consistent along the length of the implant. The quality of tissue differed greatly macroscopically between tissue produced around pulsing and non-pulsing scaffolds, but microscopically the structure of both tissues was not significantly different. Approximately 90% of cells in this tissue stained positively for CD45. Tissue in pulsing devices produced a higher amount of vimentin expression in CD45 positive staining cells than tissue in non-pulsing devices. Mechanical properties of tissue in pulsed devices were also much greater than tissue in non-pulsed devices. Two of the pulsed tissues were grafted into the carotid artery of sheep as arterial patches. In one animal tissue lasted a period of 1 week before it ruptured. In the second animal tissue lasted a period of 2 weeks at which time the animal was sacrificed. In this sheep a layer of endothelial cells had migrated to populate areas of the tissue patch. Pulsation of the implant device enhanced the development of tissue capsule in the peritoneal cavity towards arterial properties. These studies provide information on the materials and designs required to produce peritoneal-derived tissue capsules that can be used in a range of surgical applications. These studies also provide information on how this tissue responds to mechanical force and provides an in vitro system for testing this tissue. This work in this thesis has produced a device that is in the stage of pre-clinical development to be used as a potential therapy for cardiovascular disease. This device is a novel development from previous devices used for generating tissue capsules for engraftment and is a significant contribution to work in developing a replacement artery.
Keyword Bioreactor
Tissue Capsule
Tissue Engineering
Foreign Body
Additional Notes 42, 45, 50, 51, 53, 55, 58, 63, 71, 95, 99, 100, 103, 110,112, 114 to 116, 125 to 129, 131, 133, 135, 137, 143, 146, 147, 149, 151, 153 to 158, 168, 171, 176, 180 to 183, 186, 189 to 194, 196 to 200, 202, 204 to 207, 209 to 212, 214 215

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Created: Mon, 11 Jan 2010, 20:40:06 EST by Peter Stickler on behalf of Library - Information Access Service